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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2021.tde-15122022-112219
Document
Author
Full name
Tatiana Emy Koike
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2021
Supervisor
Committee
Miyabara, Elen Haruka (President)
Ferreira, Cecilia Helena de Azevedo Gouveia
Osako, Mariana Kiomy
Uchida, Marco Carlos
Title in Portuguese
Influencia do adrenoceptor β2 na funcao das celulas satelite.
Keywords in Portuguese
adrenoceptor β2
celulas satelite
Notch
proliferacao e diferenciacao
Wnt
Abstract in Portuguese
A regeneracao do musculo esqueletico e um processo altamente orquestrado que envolve a ativacao e proliferacao de celulas tronco do musculo esqueletico, denominadas celulas satelite. Uma vez ativadas, estas celulas tem a funcao de retornar ao estado quiescente ou de gerar mioblastos que se comprometem com a diferenciacao e se fundem para formar miotubos multinucleados. A sinalizacao β2-adrenergica tem se mostrado importante para o processo de regeneracao muscular, contudo o seu papel na funcao da células satélite ainda nao e bem conhecido. Diante do exposto, o objetivo deste trabalho e estudar a influencia do adrenoceptor β2 (Adrβ2) na funcao das células satélite durante a regeneracao muscular, mais especificamente analisando o papel desse receptor na autorrenovacao e proliferacao de células satélite, e na diferenciacao de celulas precursoras miogenicas utilizando-se animais knockout para o Adrβ2 (β2ko). Os resultados de experimentos in vivo mostram que os musculos tibialis anterior (TA) do grupo β2ko apresentaram significativa reducao do numero de células satélite em proliferacao (avaliadas em 3 dias apos lesao), de células satélite autorrenovadas (avaliadas em 30 dias apos lesao) e de fibras musculares em regeneracao MHCe positivas (avaliadas em 10 dias apos lesao). Nossos experimentos de proliferação in vitro evidenciam aparente menor quantidade de mioblastos em proliferação do grupo β2ko, acompanhado do aumento da expressao genica de elementos da via Notch (Notch1, Delta-like1 e RBPJ) e do inibidor do ciclo celular Cdkn1a, bem como da atividade da via Notch e redução da expressão do miR-326-3p em mioblastos em proliferacao deste grupo. Os experimentos de diferenciação muscular in vitro mostraram menor número de núcleos miogenina positivos em fibras musculares em meio de diferenciação e menor indice de fusao de mioblastos, reducao da expressao genica de elementos da via Akt/mTOR (Akt e p70S6k) e Wnt/ β-catenina (β-catenina e Lef1), queda da expressão proteica de p70S6k (ser371) e de β-catenina, redução da atividade da via Wnt/ β-catenina, além de diminuicao da expressao dos miRNAs 326-3p e 374b-5p em mioblastos em diferenciacao do grupo β2ko. Diante do exposto, nossos dados mostram que o Adrβ2 influencia a autorrenovacao das células satélite, a proliferacao de celulas precursoras miogenicas mediada pela sinalização dependente de Notch e pela expressão do miRNA 326 e, a diferenciacao de mioblastos mediada pela sinalização Wnt e pela expressão dos miRNAs 374b e 326.
Title in English
The influence of the β2 adrenoceptor in satellite cell function.
Keywords in English
β2 Adrenoceptor
Notch. Wnt
proliferation and differentiation
satellite cell
Abstract in English
Skeletal muscle regeneration is a highly orchestrated process that involves the activation and proliferation of muscle stem cells known as satellite cells. Proliferating satellite cellsmyogenic precursor cellsdifferentiate and fuse with each other, leading to the formation of multinucleated myotubes. A subpopulation of activated satellite cells self-renews to replenish the satellite cell pool. β2-adrenergic signaling is implicated in muscle regeneration; however, its role in satellite cell function is unclear. This study investigates the role of the β2 adrenoceptor (Adrβ2) in satellite cell self-renewal and proliferation and myoblast differentiation using Adrβ2 knockout (β2ko) mice. The in vitro results showed a significant reduction in the number of proliferating satellite cells (at 3 days post-injury [dpi]), self-renewed satellite cells (at 30 dpi), and regenerating eMyHC+ myofibers (at 10 dpi) in the regenerating muscle of β2ko mice. The in vitro results demonstrated a lower number of proliferating myoblasts, an increase in the gene expression of the cell cycle inhibitor Cdkn1a and Notch pathway components (Notch1, Delta-like1, and RBPJ), an increase in Notch signaling, and a decrease in the expression of miR-326-3p in proliferating myoblasts from β2ko mice. There was a decrease in the number of myogenin-positive nuclei in the differentiating myofibers, and a lower fusion index in differentiating myoblasts from β2ko mice. Furthermore, there was a decrease in the gene expression of Akt/mTOR components (Akt and p70S6k) and Wnt/ β-catenin components (β-catenin and Lef1), decreased expression of proteins p70S6k (ser371) and β-catenin, reduced activity of the Wnt/β-catenin signaling pathway, and decreased expression of miRNAs 326-3p and 374b-5p in differentiating myoblasts from β2ko mice. These results suggest that Adrβ2 influences satellite cell self-renewal and myoblast proliferation mediated by Notch signaling and miR-326-3p expression, and myoblast differentiation.
 
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Release Date
2024-12-14
Publishing Date
2022-12-22
 
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